Embryonic stem cells (ESCs) were first cultured from mouse embryos using a feeder layer of mouse fibroblasts or media conditioned with buffalo rat liver cells. The established ESC lines from mouse embryos have a characteristic phenotype consisting of a large nucleus, a prominent nucleolus, and relatively little cytoplasm. Such cells can be grown relatively indefinitely using the appropriate culture conditions. They can be induced to differentiate in vitro using retinoic acid or spontaneously by removal of the feeder layer or conditioned media. In addition, these cells can be injected into a mouse blastocyst to form a somatic and germ line chimera. This latter property has allowed mouse ESCs to be used for the production of transgenic mice with specific changes to the genome. See M. Evans et al., Nature 292, 154 (1981); G. Martin, Proc. Natl. Acad. Sci. USA 78, 7638 (1981); A. Smith et al., Developmental Biology 121, 1 (1987); T. Doetschman et al., Developmental Biology 127, 224 (1988); A. Handyside et al., Roux's Arch Dev. Biol. 198, 48 (1989).
The active compound that allows the culture of murine embryonic stem cells has been identified as differentiation inhibiting activity (DIA), also known as leukemia inhibitory factor (LIF). See A. Smith, J. Tiss. Cult. Meth. 13, 89 (1991); J. Nichols et al., Development 110, 1341 (1990). Recombinant forms of LIF can be used to obtain ESCs from mouse embryos. See S. Pease et al., Developmental Biology 141, 344 (1990).
Subsequent to the work with mouse embryos, several groups have attempted to develop stem cell lines from sheep, pig and cow. A few reports indicate that a cell line with a stem cell-like appearance has been cultured from porcine embryos using culture conditions similar to that used for the mouse. See M. Evans et al., PCT Application WO90/03432; E. Notarianni et al., J. Reprod. Fert., Suppl. 41, 51 (1990); J. Piedrahita et al., Theriogenology 34, 879 (1990); E. Notarvianni et al., Proceedings of the 4th World Congress on Genetics Applied to Livestock Production, 58 (Edinburgh, July 1990).
Few or no attempts have been made to date regarding the culture of embryonic stem cells from avian embryos. The main reason for this is that it is very difficult to establish a continuous line of chicken cells without viral or chemical transformation, and most primary chicken lines do not survive beyond 2-3 months. The culture of cells from the unincubated embryo has been more difficult, and under reported conditions such cells do not survive beyond two weeks. See E. Mitrani et al., Differentiation 21, 56-61 (1982); E. Sanders et al., Cell Tissue Res. 220, 539 (1981).